The immune system uses many mechanisms for attacking pathogens; however, not all of these mechanisms are necessarily activated after immunization. Protective immunity induced by vaccination is dependent on the capacity of the vaccine to elicit the appropriate immune response to resist or eliminate the pathogen. Depending on the pathogen, this may require a cell-mediated and/or humoral immune response.
The current paradigm for the role of helper T cells in the immune response is that T cells can be separated into subsets on the basis of the cytokines they produce, and that the distinct cytokine profile observed in these cells determines their function. This T cell model Includes two major subsets: TH-1 cells that produce IL-2 and interferon xcex3 (IFN-xcex3) which augment both cellular and humoral immune responses, and TH-2 cells that produce IL-4, IL-5 and IL-10 which augment humoral immune responses (Mosmann et al., J. Immunol. 126:2348 (1986)). It is often desirable to enhance the immunogenic potency of an antigen in order to obtain a stronger immune response in the organism being immunized and to strengthen host resistance to the antigen-bearing agent. A substance that enhances the immunogenicity of an antigen with which it is administered is known as an adjuvant. For example, certain lymphokines have been shown to have adjuvant activity, thereby enhancing the immune response to an antigen (Nencioni et al., J. Immunol. 139:800-804 (1987); EP285441 to Howard et al.).
This invention pertains to vaccine compositions comprising a mixture of herpes simplex virus glycoprotein D, the interleukin IL-12 and a mineral in suspension. The IL-12 may be either adsorbed onto the mineral suspension or simply mixed therewith. In a particular embodiment of the invention, the IL-12 is adsorbed onto a mineral suspension such as alum (e.g., aluminum hydroxide or aluminum phosphate). In a particular embodiment, the IL-12 is human IL-12. The invention also pertains to vaccine compositions which further comprise a physiologically acceptable vehicle. The invention further relates to immunogenic compositions comprising a mixture of a herpes simplex virus glycoprotein D, an adjuvant amount of interleukin-12, a mineral in suspension, and optionally comprising a physiologically acceptable vehicle.
The compositions of the present invention modulate the protective immune response to the antigen; that is, the vaccine composition is capable of quantitatively and qualitatively improving the vaccinated host""s antibody response, and quantitatively increasing cell-mediated immunity for a protective response to a pathogen. In a particular embodiment of the invention, the antigen is a herpes simplex viral (HSV) antigen, such as envelope glycoprotein D (gD) of herpes simplex virus types I and/or II.
The invention also pertains to methods for preparing a vaccine composition comprising mixing HSV gD and IL-12 with a mineral in suspension. In particular, the IL-12 is adsorbed onto the mineral suspension. The invention also pertains to methods for eliciting or increasing a vaccinate""s humoral and/or cell-mediated immunity, for a protective immune response, comprising administering to a vertebrate host an effective amount of a vaccine composition comprising a mixture of HSV gD, IL-12 and a mineral in suspension in a physiologically acceptable solution. In particular, the IL-12 is adsorbed onto the mineral suspension.
Glycoprotein D (gD) is an envelope glycoprotein of herpes simplex virus (HSV) types I and II. HSV gD has been shown to be a potent inducer of protective immunity against primary and recurrent HSV infection in animal models (Mishkin et al., Vaccine 9:147-153 (1991); Landolfi et al., Vaccine 11:407-414 (1993)).
IL-12 is produced by a variety of antigen-presenting cells, principally macrophages and monocytes. It is a critical element in the induction of TH-1 cells from naive T cells. Production of IL-12 or the ability to respond to it has been shown to be critical in the development of protective TH-1-like responses, for example, during parasitic infections, most notably Leishmaniasis (Scott et al., U.S. Pat. No. 5,571,515). The effects of IL-12 are mediated by IFN-xcex3 produced by NK cells and T helper cells. IFN-xcex3 is critical for the induction of IgG2a antibodies to T-dependent protein antigens (Finkelman and Holmes, Annu. Rev. Immunol. 8:303-33 (1990)) and IgG3 responses to T-independent antigens (Snapper et al., J. Exp. Med. 175:1367-1371 (1992)). Interleukin-12 (IL-12), originally called natural killer cell stimulatory factor, is a heterodimeric cytokine (Kobayashi et al., J. Exp. Med. 170:827 (1989)). The expression and isolation of IL-12 protein in recombinant host cells is described in International Patent Application WO 90/05147, published May 17, 1990.
The studies described herein relate to the utility of IL-12 as an adjuvant in a herpes simplex virus (HSV) vaccine. Accordingly, this invention pertains to vaccine compositions comprising a mixture of HSV gD, IL-12 and a mineral in suspension. In a particular embodiment of the invention, the IL-12 is adsorbed onto a mineral suspension such as alum (e.g., aluminum hydroxide or aluminum phosphate). These vaccine compositions modulate the protective immune response to HSV; that is, the vaccine composition is capable of eliciting the vaccinated host""s cell-mediated immunity for a protective response to the pathogenic antigen.
IL-12 can be obtained from several suitable sources. It can be produced by recombinant DNA methodology; for example, the gene encoding human IL-12 has been cloned and expressed in host systems, permitting the production of large quantities of pure human IL-12. Also useful in the present invention are biologically active subunits or fragments of IL-12. Further, certain T lymphocyte lines produce high levels of IL-12, thus providing a readily available source. Commercial sources of recombinant human and murine IL-12 include Genetics Institute, Inc. (Cambridge, Mass.).
The antigen of this invention, e.g., an HSV antigen, can be used to elicit an immune response to she antigen in a vertebrate such as a mammalian host. For example, the antigen can be an HSV gD protein antigen or a portion thereof which retains the ability to stimulate an immune response.
The method of the present invention comprises administering to a mammal, particularly a human or other primate, an immunologically effective dose of a vaccine composition comprising a mixture of an antigen, e.g., an HSV gD antigen, an adjuvant amount of IL-12 and a mineral in suspension. In particular, the IL-12 is adsorbed onto the mineral suspension. As used herein, an xe2x80x9cadjuvant amountxe2x80x9d of IL-12 is intended to mean doses of from about 1 nanogram to about 20 micrograms, and more particularly from about 100 nanograms to about 5 micrograms. As used herein, an xe2x80x9cimmunologically effectivexe2x80x9d dose of the vaccine composition is a dose which is suitable to elicit an immune response. The particular dosage will depend upon the age, weight and medical condition of the mammal to be treated, as well as on the method of administration. Suitable doses will be readily determined by the skilled artisan. The vaccine composition can be optionally administered in a pharmaceutically or physiologically acceptable vehicle, such as physiological or phosphate buffered saline or ethanol polyols such as glycerol or propylene glycol. A small amount of detergent may also be included to enhance vaccine stability.
The vaccine composition may optionally comprise additional adjuvants such as vegetable oils or emulsions thereof, surface active substances, e.g., hexadecylamin, octadecyl amino acid esters, octadecylamine, lysolecithin, dimethyl-dioctadecylammonium bromide, N,N-dicoctadecyl-Nxe2x80x2-Nxe2x80x2bis(2-hydroxyethyl-propane diamine), methoxyhexadecylglycerol, and pluronic polyols; polyamines, e.g., pyran, dextransulfate, poly IC, carbopol; peptides, e g., muramly. dipeptide, dimethylglycine, tuftsin; immune stimulating complexes; oil emulsions; lipopolysaccharides such as MPL (3-O-deacylated monophosphoryl lipid A; RIBI ImmunoChem Research, Inc., Hamilton, Mont.); and mineral gels. The antigens of this invention can also be incorporated into liposomes, cochleates, biodegradable polymers such as poly-lactide, poly-glycolide and poly-lactide-co-glycolides, or ISCOMS (immunostimulating complexes), and supplementary active ingredients may also be employed. Antigens of the present invention can also be administered in combination with bacterial toxins and their attenuated derivatives. The antigens of the invention can also be administered in combination with other lymphokines including, but not limited to, interleukin-2, IFN-xcex3 and GM-CSF.
The vaccines can be administered to a human or animal by a variety of routes, including but not limited to parenteral, intraarterial, intradermal, transdermal (such as by the use of slow release polymers), intramuscular, intraperitoneal, intravenous, subcutaneous, oral and intranasal routes of administration. The amount of antigen employed in such vaccines will vary depending upon the identity of the antigen. Adjustment and manipulation of established dosage ranges used with traditional carrier antigens for adaptation to the present vaccine is well within the ability of those skilled in the art. The vaccines of the present invention are intended for use in the treatment of both immature and adult warm-blooded animals, and, in particular, humans. Typically, the antigen and IL-12/alum combination will be administered at the same time.
The adjuvant action of IL-12 has a number of important implications. The adjuvanticity of IL-12 can increase the concentration of protective antibodies produced against the antigen in the vaccinated organism. As a result, effective (i.e., protective) vaccination can be achieved with a smaller quantity of antigen than would be normally required. This reduction in the required amount of antigen may lead to more widespread use of vaccines which are difficult and costly to prepare. Additionally, the use of IL-12 as an adjuvant can enhance the ability of antigens which are weakly antigenic or poorly immunogenic to elicit an immune response. It may also provide for safer vaccination when the antigen is toxic at the concentration normally required for effective immunization. By reducing the amount of antigen, the risk of toxic reaction is reduced.
Typically, vaccination regimens call for the administration of antigen over a period of weeks or months in order to stimulate a xe2x80x9cprotectivexe2x80x9d immune response. A protective immune response is an immune response sufficient to protect the immunized organism from disease caused by a particular pathogen or pathogens to which the vaccine is directed. IL-12, when administered with an antigen, such as an HSV antigen, and mixed with or adsorbed onto a mineral (e.g., alum) in suspension can accelerate the generation of a protective immune response. This may reduce the time course of effective vaccination regimens. In some instances, it may result in the generation of a protective response in a single dose. The vaccine compositions of this invention are also useful therapeutically, to reduce the number and severity of symptomatic episodes in subjects already infected with HSV.
As the result of work described herein, coadministration of HSV subunit vaccine with IL-12 adsorbed on alum in suspension has been shown to elicit an overwhelmingly TH-1-associated profile of response; this is a novel pattern of immune induction for gD subunit vaccine. As further described herein, dose ranges of TL-12 operative in preclinical models of immunization with soluble and aluminum phosphate adsorbed gD have been determined Results described herein also reveal that immunization with the cytokine/glycoprotein combination with or without alum elicits a TH-1-associated antibody profile. Coadministration of vaccine and IL-12/alum has adjuvant effects on immune responsiveness to the subunit vaccine as indicated by increases in the level of anti-gD antibodies measured by ELISA and virus neutralization.
Glycoprotein D (gD), an envelope glycoprotein of herpes simplex virus types I and II shown to be requisite for virus infectivity and a major target of humoral and cellular immune responses, serves as a primary vaccine candidate for use against primary and recurrent herpes infection in humans. Administration of gD subunit vaccine formulated with aluminum-based or other immunoadsorbents currently accepted for use in humans has been shown in several preclinical studies to induce a profile of immune response attributable to a predominant stimulation of TH-2 T helper lymphocytes. However, it appears that interdiction of recurrent herpes disease and the establishment of appropriate protective immune function will require the induction of a potent TH-1 profile of response.
Results of work described herein indicate that coadministration of IL-12 with soluble and AlPO4-adsorbed gD results in a quantitative increase in humoral and cellular immune responses, as well as in a Qualitative alteration in the humoral response. This is demonstrated by the altered IgG subclass profile elicited by the vaccine in the presence of IL-12. Indeed, the preferential induction of IgG2a antibodies, with their efficient complement fixing capability, is one of the hallmarks of a TH-1 response profile. The immunomodulation of the immune response by IL-12 administration is also evident in the shifted proportion and magnitude of IFN-xcex3 (TH-1 associated) secretion when compared with IL-4 (TH-2 associated).
Furthermore, it is particularly important to note the induction of antigen-specific cytolytic activity in mice immunized with soluble subunit vaccine coadministered with IL-12. This pattern of response suggests that the addition of IL-12 results in a fundamental change in the character of the immune response to gD subunit vaccines, because gD subunit vaccines have rarely, if ever, demonstrated the ability to induce cytolytic activity in previous studies.
The spectrum of immune response elicited by this novel formulation is closely correlated with that induced by natural virus infection and associated with a pattern of immunity observed in disease free seropositive humans. Taken together, these results suggest that IL-12-mediated immunomodulation provides a significant benefit in establishing an immunological response profile effective in immunotherapeutic and/or prophylactic intervention against herpes simplex virus disease.
The following Examples are offered for the purpose of illustrating the present invention and are not to be construed to limit the scope of this invention. The teachings of all references cited herein are hereby incorporated herein by reference.